CN100340669C - 10 ton liquid submerged femrentation culturing method of Gleoesterum incarnatum - Google Patents
10 ton liquid submerged femrentation culturing method of Gleoesterum incarnatum Download PDFInfo
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Abstract
The present invention discloses a submerged fermentation culture method of ten tons of glue tenacity leather fungus liquid, which belongs to the field of edible fungus fermentation industry, particularly to a submerged fermentation culture method of the glue tenacity leather fungus liquid. The submerged fermentation culture method is characterized in that the submerged fermentation culture method comprises activation culture of inclined plane strains, culture of first grade strains, culture of second grade strains, culture of third grade strains, tank culture of the first grade strains, tank culture of the second grade strains and fermentation tank culture. A carbon source for the stains and fermentation culture comprises general raw material, such as glucose, sucrose, starch, etc.; a nitrogen source comprises peptone, defatted soybean protein powder, bean cake powder, ammonium sulfate, etc.; inorganic salt which is required to be added comprises potassium dihydrogen phosphate, magnesium sulfate and vitamin B1. The submerged fermentation culture method determines the submerged fermentation culture method of ten tons of glue tenacity leather fungus liquid and lays a solid foundation for developing and processing relevant products of glue tenacity leather fungi.
Description
Technical field
The invention belongs to edible fungus fermented industrial circle, particularly relate to the gleoesterum incarnatum liquid submerged femrentation culturing method.
Background technology
Gleoesterum incarnatum [Gloeostereum incarnatum], another name elm ear, elm mill.Belong to Basidiomycotina, Hymenomycetes, Aphyllophorales, photovoltaicing leather bacteria section, gleoesterum incarnatum genus.Wild gleoesterum incarnatum mainly is distributed in China the Northeast and Hokkaido, Japan, the Tieling in Liaoning Province, former clearly, new guest, Benxi and Fushun, the Tonghua in Jilin Province, Huijiang, Fusong, long white and Antu etc., the Eastern Mountain Area of Heilongjiang Province is the main producing region of elm ear, also there is a small amount of distribution in Xinjiang.Gleoesterum incarnatum is famous dietotherapeutic fungi.The elm ear is introduced the existing long history of Chinese medicine.The famous medical scholar's LI Shi-Zhen of the Ming Dynasty is shown in the Compendium of Material Medica and to be carried: " the elm ear is adopted it August ", " August, the elm ear exposed to the sun with the good wine stain, and same foxtail millet seed, real the cooking of purple amaranth are the end.Every clothes a dose of powder just as much as the amount can be taken up with three fingers, under the wine, it is not hungry to make us warding off paddy." gleoesterum incarnatum sporophore nutritive ingredient is very abundant.Analyze according to Chinese Academy Of Preventive Medicine Research Institute Of Nutrition And Food Hygiene, gleoesterum incarnatum mainly contains protein, carbohydrate, vitamin-E, vitamins B
1, vitamins B
2, necessary each seed amino acid of human body such as trace element such as calcium, phosphorus, iron, zinc and L-glutamic acid, Methionin.The content of various functional components and sporophore are basic identical in the mycelium of fermentative production.
Folks of china is treated enteritis often behind the gleoesterum incarnatum decocting.Luan Hengchun, Tai Longjie etc. have delivered the article of " gleoesterum incarnatum (Gloeostereumincarnatum S.Itoet Imal) research (I)---the research of the external bacteriostatic action of tunning " at 1994 the 2nd phase Northeast China Normal University's journals.Paper has been set forth the obvious restraining effect of gleoesterum incarnatum fermented liquid to the human pathogen.
The gleoesterum incarnatum wild resource is extremely rare.It is two kinds of effective ways that obtain gleoesterum incarnatum that artificial culture and liquid submerged fermentation are cultivated, but artificial cultivation technique is still immature at present, does not possess the mature technology of implant mass.It is a kind of effective ways that can obtain gleoesterum incarnatum in a large number that liquid submerged fermentation is cultivated, but has only laboratory scale gleoesterum incarnatum liquid submerged fermentation culture technique at present, the deep liquid culture technique still is in testing laboratory's level, Tai Longjie, Luan Hengchun, Zhao Qiang etc. have delivered the article of " gleoesterum incarnatum (Gloeostereumincarnatum S.Itoet Imal) research II---deep layer is cultivated " at 1997 the 3rd phase Northeast China Normal University's journals, and the laboratory scale fermentative Production method of gleoesterum incarnatum reported in article.But the heavy industrialization deep liquid is cultivated the method for gleoesterum incarnatum and is not seen disclosure as yet.Because industrially scalable deep liquid cultural method is subjected to the influence of fermentation equipment and processing parameter, its cultural method need be furtherd investigate and test just and can obtain.The shortage of large scale culturing gleoesterum incarnatum method has influenced the further development and use of gleoesterum incarnatum good efficacy.
Summary of the invention:
The present invention has overcome the defective that lacks the heavy industrialization cultural method in the present gleoesterum incarnatum cultivation, and the technical problem that the present invention solves provides a kind of extensive (10 tons) gleoesterum incarnatum deep liquid cultural method.
Technical scheme of the present invention is summarized as follows:
(1) slant strains activation culture: the gleoesterum incarnatum slant strains of preservation is transferred on the slant medium, cultivated about 96～120 hours, and covered with the inclined-plane to mycelia and get final product for 22～28 ℃; 25 ℃ of optimum culturing temperatures.
(2) first order seed is cultivated: access of above-mentioned gleoesterum incarnatum slant strains picking is equipped with carries out first order seed in 500 ml shake flasks of 100 milliliters of substratum and cultivate, culture condition: 80～180 rev/mins of rotary shaking tables, cultivated about 96～130 hours for 22～28 ℃; Rotary shaking table is advisable for 150 rev/mins, and culture temperature is advisable with 25 ℃.
(3) secondary seed is cultivated: the one-level shake-flask seed is inserted with 5～20% inoculum sizes carry out the secondary seed cultivation in 500 ml shake flasks that 100 milliliters of substratum are housed, culture condition: 80～180 rev/mins of rotary shaking tables, cultivated about 96～130 hours for 22～28 ℃; Suitable inoculum size is 10%, and rotary shaking table is advisable for 150 rev/mins, and culture temperature is advisable with 25 ℃.
(4) three grades of seed culture: the secondary shake-flask seed inserted in 5000 ml shake flasks that 1000 milliliters of substratum are housed with 5～20% inoculum sizes carry out three grades of seed culture, culture condition: 80～180 rev/mins of rotary shaking tables, cultivated 72～96 hours for 22～28 ℃; Suitable inoculum size is 10%, and rotary shaking table is advisable for 100 rev/mins, and culture temperature is advisable with 25 ℃.
(5) first class seed pot is cultivated: the cubic capacity that the inoculum size with 5～20% is equipped with 0.1 ton of fermention medium to three grades of seeds accesses is 0.15 ton a first class seed pot.Culture condition is: 22～28 ℃ of culture temperature, and 80～180 rev/mins of stirring velocitys, ventilation (V/V) 1: 0.3～0.8, tank pressure 0.03～0.1Mpa cultivated 72～96 hours; Suitable inoculum size is 15%, 25 ℃ of optimum culturing temperatures, 120 rev/mins of best stirring velocitys, optimal ventilation amount (V/V) 1: 0.6, best tank pressure 0.05Mpa.
(6) the secondary seed jar is cultivated: is in 1.5 tons of secondary seed jars with the bacterium liquid of first class seed pot with 5～20% the inoculum size cubic capacity that 1.0 tons of fermention mediums are housed of transferring, culture condition is: 22～28 ℃ of culture temperature, 80～180 rev/mins of stirring velocitys, ventilation (V/V) 1: 0.3～0.8, tank pressure 0.03～0.1Mpa cultivated 72～96 hours; Suitable inoculum size is 15%, 25 ℃ of optimum culturing temperatures, 120 rev/mins of best stirring velocitys, optimal ventilation amount (V/V) 1: 0.6, best tank pressure 0.05Mpa.
(7) fermentor cultivation: it is in 15 tons the fermentor tank that the inoculum size with 5～20% inserts the cubic capacity that 10 tons of fermention mediums are housed with secondary seed jar seed.Culture condition is: 22～28 ℃ of culture temperature, and 80～180 rev/mins of stirring velocitys, ventilation (V/V) 1: 0.3～0.8, tank pressure 0.03～0.1Mpa cultivated 150～200 hours; Suitable inoculum size is 15%, 25 ℃ of optimum culturing temperatures, 120 rev/mins of best stirring velocitys, optimal ventilation amount (V/V) 1: 0.6, best tank pressure 0.05Mpa.
The dried mycelia yield of gleoesterum incarnatum is between 1.5%～3.0% among the present invention.
Mycelium dry weight measuring method of the present invention: with 10 minutes centrifugation fermented liquids of whizzer 3000rpm, collect wet mycelium, use sterilized water centrifuge washing 2 times, it promptly is mycelium dry weight that wet mycelium is weighed in 60-80 ℃ of freeze-day with constant temperature to constant weight.
Mycelium weight in wet base measuring method of the present invention:, collect wet mycelium and weigh promptly with 10 minutes centrifugation fermented liquids of whizzer 3000rpm.
The gleoesterum incarnatum bacterial classification is purchased in Chinese industrial microbial strains preservation administrative center among the present invention, bacterium number: CICC14024.
The slant medium composition can be PDA slant medium or other suitable culture mediums among the present invention.
Seed and fermentation culture carbon source can be selected raw materials commonly used such as glucose, sucrose, starch among the present invention, and nitrogenous source can be selected peptone, defatted soybean protein powder, soybean cake powder, ammonium sulfate etc.; The inorganic salt that need to add have potassium primary phosphate, sal epsom, and VITAMIN is added VITMAIN B1, and substratum initial p H is controlled at about 6.0～7.0.
Seed culture medium is identical with the fermention medium composition, and its ratio is: starch 2～3%, sucrose 3～5%, glucose 1～3%, peptone 0.5%, defatted soybean protein powder 2～3%, potassium primary phosphate 0.13～0.2%, sal epsom 0.1～0.15%, vitamins B
12PPm, PH6～7.
Putting jar condition among the present invention is: wet mycelium content is more than 20%, and fermented liquid is by rare retrogradation, and by muddy bleach, pH value about 3.5～4.5 gets final product.Fermented liquid is operated after filtration and can be obtained fermentation clear liquid and mycelium.
The present invention has determined 10 tons of fermentation broth liquor submerged culturing method for making of gleoesterum incarnatum by the repetition test of substratum and culture condition, and the dried mycelia yield of gleoesterum incarnatum is between 1.5%～3.0%.Invention has obtained good effect.For exploitation processing gleoesterum incarnatum related products is had laid a good foundation.
The inventive method is applicable to that gleoesterum incarnatum belongs to other bacterial classification and carries out large scale culturing by fermentation method.
Embodiment:
The following examples can make those skilled in the art more fully understand the present invention, but do not limit the present invention in any way.
Embodiment 1 (10 tons of fermented liquids)
(1) slant strains activation culture: the gleoesterum incarnatum slant strains of preservation is transferred on the slant medium, cultivated 120 hours for 25 ℃, mycelia is covered with the inclined-plane;
(2) first order seed is cultivated: access of above-mentioned gleoesterum incarnatum slant strains picking is equipped with carries out first order seed in 500 ml shake flasks of 100 milliliters of substratum and cultivate culture condition: 80 rev/mins of rotary shaking tables, cultivated 130 hours for 25 ℃.
(3) secondary seed is cultivated: the one-level shake-flask seed is inserted with 5% inoculum size carry out the secondary seed cultivation in 500 ml shake flasks that 100 milliliters of substratum are housed, culture condition: 80 rev/mins of rotary shaking tables, cultivated 120 hours for 25 ℃.
(4) three grades of seed culture: the secondary shake-flask seed inserted in 5000 ml shake flasks that 1000 milliliters of substratum are housed with 5% inoculum size carry out three grades of seed culture, culture condition: 80 rev/mins of rotary shaking tables, cultivated 96 hours for 25 ℃.
(5) first class seed pot is cultivated: the cubic capacity that the inoculum size with 5% is equipped with 0.1 ton of fermention medium to three grades of seeds accesses is 0.15 ton a first class seed pot.Culture condition is: 25 ℃ of culture temperature, and 80 rev/mins of stirring velocitys, ventilation (V/V) 1: 0.5, tank pressure 0.03Mpa cultivated 96 hours.
(6) the secondary seed jar is cultivated: is in 1.5 tons of secondary seed jars with the bacterium liquid of first class seed pot with 5% the inoculum size cubic capacity that 1.0 tons of fermention mediums are housed of transferring, culture condition is: 25 ℃ of culture temperature, 80 rev/mins of stirring velocitys, ventilation (V/V) 1: 0.5, tank pressure 0.03Mpa cultivated 96 hours.
(7) fermentor cultivation: it is in 15 tons the fermentor tank that the inoculum size with 5% inserts the cubic capacity that 10 tons of fermention mediums are housed with secondary seed jar seed.Culture condition is: 25 ℃ of culture temperature, and 80 rev/mins of stirring velocitys, ventilation (V/V) 1: 0.3, tank pressure 0.03Mpa cultivated 200 hours.
Slant medium consists of the PDA slant medium in this example.
Seed culture medium and fermention medium consist of in this example: starch 2%, sucrose 4%, glucose 2%, peptone 0.5%, defatted soybean protein powder 2.0%, potassium primary phosphate 0.2%, sal epsom 0.1%, vitamins B
12PPm, PH6.0.
The dried mycelia yield of gleoesterum incarnatum is 2.5% in this example.
Embodiment 2 (10 tons of fermented liquids)
(1) slant strains activation culture: the gleoesterum incarnatum slant strains of preservation is transferred on the slant medium, cultivated 120 hours for 22 ℃, mycelia is covered with the inclined-plane.
(2) first order seed is cultivated: access of above-mentioned gleoesterum incarnatum slant strains picking is equipped with carries out first order seed in 500 ml shake flasks of 100 milliliters of substratum and cultivate culture condition: 150 rev/mins of rotary shaking tables, cultivated 130 hours for 25 ℃.
(3) secondary seed is cultivated: the secondary shake-flask seed is inserted with 10% inoculum size carry out the secondary seed cultivation in 500 ml shake flasks that 100 milliliters of substratum are housed, culture condition: 100 rev/mins of rotary shaking tables, cultivated 100 hours for 25 ℃.
(4) three grades of seed culture: the secondary shake-flask seed inserted in 5000 ml shake flasks that 1000 milliliters of substratum are housed with 10% inoculum size carry out three grades of seed culture, culture condition: 100 rev/mins of rotary shaking tables, cultivated 72 hours for 25 ℃.
(5) first class seed pot is cultivated: the cubic capacity that the inoculum size with 10% is equipped with 0.1 ton of fermention medium to three grades of seeds accesses is 0.15 ton a first class seed pot.Culture condition is: 25 ℃ of culture temperature, and 100 rev/mins of stirring velocitys, ventilation (V/V) 1: 0.5, tank pressure 0.05Mpa cultivated 80 hours.
(6) the secondary seed jar is cultivated: is in 1.5 tons of secondary seed jars with the bacterium liquid of first class seed pot with 10% the inoculum size cubic capacity that 1.0 tons of fermention mediums are housed of transferring, culture condition is: 25 ℃ of culture temperature, 100 rev/mins of stirring velocitys, ventilation (V/V) 1: 0.3, tank pressure 0.05Mpa cultivated 80 hours.
(7) fermentor cultivation: it is in 15 tons the fermentor tank that the inoculum size with 10% inserts the cubic capacity that 10 tons of fermention mediums are housed with secondary seed jar seed.Culture condition is: 25 ℃ of culture temperature, and 100 rev/mins of stirring velocitys, ventilation (V/V) 1: 0.5, tank pressure 0.05Mpa cultivated 180 hours.
Slant medium consists of the PDA slant medium in this example.
Seed culture medium and fermention medium consist of in this example: starch 2%, sucrose 3%, glucose 2%, peptone 0.5%, defatted soybean protein powder 2.0%, potassium primary phosphate 0.2%, sal epsom 0.1%, vitamins B
12PPm, PH6.0.
The dried mycelia yield of gleoesterum incarnatum is 1.5% in this example.
Embodiment 3 (10 tons of fermented liquids)
(1) slant strains activation culture: the gleoesterum incarnatum slant strains of preservation is transferred on the slant medium, cultivated 100 hours for 25 ℃, mycelia is covered with the inclined-plane;
(2) first order seed is cultivated: access of above-mentioned gleoesterum incarnatum slant strains picking is equipped with carries out first order seed in 500 ml shake flasks of 100 milliliters of substratum and cultivate culture condition: 120 rev/mins of rotary shaking tables, cultivated 100 hours for 25 ℃.
(3) secondary seed is cultivated: the one-level shake-flask seed is inserted with 15% inoculum size carry out the secondary seed cultivation in 500 ml shake flasks that 100 milliliters of substratum are housed, culture condition: 150 rev/mins of rotary shaking tables, cultivated 96 hours for 25 ℃.
(4) three grades of seed culture: the secondary shake-flask seed inserted in 5000 ml shake flasks that 1000 milliliters of substratum are housed with 15% inoculum size carry out three grades of seed culture, culture condition: 100 rev/mins of rotary shaking tables, cultivated 96 hours for 25 ℃.
(5) first class seed pot is cultivated: the cubic capacity that the inoculum size with 15% is equipped with 0.1 ton of fermention medium to three grades of seeds accesses is 0.15 ton a first class seed pot.Culture condition is: 24 ℃ of culture temperature, and 120 rev/mins of stirring velocitys, ventilation (V/V) 1: 0.5, tank pressure 0.03Mpa cultivated 96 hours.
(6) the secondary seed jar is cultivated: is in 1.5 tons of secondary seed jars with the bacterium liquid of first class seed pot with 15% the inoculum size cubic capacity that 1.0 tons of fermention mediums are housed of transferring, culture condition is: 25 ℃ of culture temperature, 120 rev/mins of stirring velocitys, ventilation (V/V) 1: 0.5, tank pressure 0.03Mpa cultivated 96 hours.
(7) fermentor cultivation: it is in 15 tons the fermentor tank that the inoculum size with 15% inserts the cubic capacity that 10 tons of fermention mediums are housed with secondary seed jar seed.Culture condition is: 25 ℃ of culture temperature, and 120 rev/mins of stirring velocitys, ventilation (V/V) 1: 0.3, tank pressure 0.03Mpa cultivated 160 hours.
Slant medium consists of the PDA slant medium in this example.
Seed culture medium and fermention medium consist of in this example: starch 3%, sucrose 3%, glucose 2%, peptone 0.5%, defatted soybean protein powder 2.0%, potassium primary phosphate 0.15%, sal epsom 0.1%, vitamins B
12PPm, PH6.0.
The dried mycelia yield of gleoesterum incarnatum is 2.0% in this example.
Embodiment 4 (10 tons of fermented liquids)
(1) slant strains activation culture: the gleoesterum incarnatum slant strains of preservation is transferred on the slant medium, cultivated about 96 hours, and covered with the inclined-plane to mycelia and get final product for 28 ℃; 28 ℃ of optimum culturing temperatures.
(2) first order seed is cultivated: access of above-mentioned gleoesterum incarnatum slant strains picking is equipped with carries out first order seed in 500 ml shake flasks of 100 milliliters of substratum and cultivate culture condition: 180 rev/mins of rotary shaking tables, cultivated about 96 hours for 28 ℃.
(3) secondary seed is cultivated: the one-level shake-flask seed is inserted with 20% inoculum size carry out the secondary seed cultivation in 500 ml shake flasks that 100 milliliters of substratum are housed, culture condition: 80 rev/mins of rotary shaking tables, cultivated about 96 hours for 28 ℃.
(4) three grades of seed culture: the secondary shake-flask seed inserted in 5000 ml shake flasks that 1000 milliliters of substratum are housed with 20% inoculum size carry out three grades of seed culture, culture condition: 180 rev/mins of rotary shaking tables, cultivated 72 hours for 28 ℃.
(5) first class seed pot is cultivated: the cubic capacity that the inoculum size with 20% is equipped with 0.1 ton of fermention medium to three grades of seeds accesses is 0.15 ton a first class seed pot.Culture condition is: 28 ℃ of culture temperature, and 180 rev/mins of stirring velocitys, ventilation (V/V) 1: 0.8, tank pressure 0.1Mpa cultivated 72 hours.
(6) the secondary seed jar is cultivated: is in 1.5 tons of secondary seed jars with the bacterium liquid of first class seed pot with 20% the inoculum size cubic capacity that 1.0 tons of fermention mediums are housed of transferring, culture condition is: 28 ℃ of culture temperature, 180 rev/mins of stirring velocitys, ventilation (V/V) 1: 0.8, tank pressure 0.1Mpa cultivated 72 hours.
(7) fermentor cultivation: it is in 15 tons the fermentor tank that the inoculum size with 20% inserts the cubic capacity that 10 tons of fermention mediums are housed with secondary seed jar seed.Culture condition is: 28 ℃ of culture temperature, and 180 rev/mins of stirring velocitys, ventilation (V/V) 1: 0.8, tank pressure 0.1Mpa cultivated 150 hours.
The slant medium composition can be the PDA slant medium in this example.
Seed culture medium and fermention medium consist of in this example: starch 2%, sucrose 4%, glucose 2%, peptone 0.5%, defatted soybean protein powder 2.0%, potassium primary phosphate 0.15%, sal epsom 0.1%, vitamins B
12PPm, PH6.0.
The dried mycelia yield of gleoesterum incarnatum is 2.4% in this example.
Embodiment 5 (10 tons of fermented liquids)
(1) slant strains activation culture: the gleoesterum incarnatum slant strains of preservation is transferred on the slant medium, cultivated about 120 hours, and covered with the inclined-plane to mycelia and get final product for 22 ℃;
(2) first order seed is cultivated: access of above-mentioned gleoesterum incarnatum slant strains picking is equipped with carries out first order seed in 500 ml shake flasks of 100 milliliters of substratum and cultivate culture condition: 150 rev/mins of rotary shaking tables, cultivated about 130 hours for 22 ℃;
(3) secondary seed is cultivated: the one-level shake-flask seed is inserted with 20% inoculum size carry out the secondary seed cultivation in 500 ml shake flasks that 100 milliliters of substratum are housed, culture condition: 100 rev/mins of rotary shaking tables, cultivated about 96 hours for 22 ℃;
(4) three grades of seed culture: the secondary shake-flask seed inserted in 5000 ml shake flasks that 1000 milliliters of substratum are housed with 20% inoculum size carry out three grades of seed culture, culture condition: 130 rev/mins of rotary shaking tables, cultivated 80 hours for 22 ℃;
(5) first class seed pot is cultivated: the cubic capacity that the inoculum size with 20% is equipped with 0.1 ton of fermention medium to three grades of seeds accesses is 0.15 ton a first class seed pot.Culture condition is: 22 ℃ of culture temperature, and 80 rev/mins of stirring velocitys, ventilation (V/V) 1: 0.8, tank pressure 0.1Mpa cultivated 80 hours;
(6) the secondary seed jar is cultivated: is in 1.5 tons of secondary seed jars with the bacterium liquid of first class seed pot with 20% the inoculum size cubic capacity that 1.0 tons of fermention mediums are housed of transferring, culture condition is: 22 ℃ of culture temperature, 80 rev/mins of stirring velocitys, ventilation (V/V) 1: 0.8, tank pressure 0.06Mpa cultivated 96 hours;
(7) fermentor cultivation: it is in 15 tons the fermentor tank that the inoculum size with 15% inserts the cubic capacity that 10 tons of fermention mediums are housed with secondary seed jar seed.Culture condition is: 22 ℃ of culture temperature, and 120 rev/mins of stirring velocitys, ventilation (V/V) 1: 0.4, tank pressure 0.05Mpa cultivated 180 hours;
The slant medium composition can be PDA slant medium or other suitable culture mediums in this example.
Seed culture medium and fermention medium consist of in this example: starch 2%, sucrose 5%, glucose 1%, peptone 0.5%, defatted soybean protein powder 3.0%, potassium primary phosphate 0.15%, sal epsom 0.15%, vitamins B
12PPm, PH7.0.
The dried mycelia yield of gleoesterum incarnatum is 1.7% in this example.
Embodiment 6 (10 tons of fermented liquids)
(1) slant strains activation culture: the gleoesterum incarnatum slant strains of preservation is transferred on the slant medium, cultivated about 96 hours, and covered with the inclined-plane to mycelia and get final product for 26 ℃.
(2) first order seed is cultivated: access of above-mentioned gleoesterum incarnatum slant strains picking is equipped with carries out first order seed in 500 ml shake flasks of 100 milliliters of substratum and cultivate culture condition: 130 rev/mins of rotary shaking tables, cultivated about 90 hours for 26 ℃;
(3) secondary seed is cultivated: the one-level shake-flask seed is inserted with 10% inoculum size carry out the secondary seed cultivation in 500 ml shake flasks that 100 milliliters of substratum are housed, culture condition: 140 rev/mins of rotary shaking tables, cultivated about 96 hours for 26 ℃;
(4) three grades of seed culture: the secondary shake-flask seed inserted in 5000 ml shake flasks that 1000 milliliters of substratum are housed with 10% inoculum size carry out three grades of seed culture, culture condition: 100 rev/mins of rotary shaking tables, cultivated 72 hours for 26 ℃;
(5) first class seed pot is cultivated: the cubic capacity that the inoculum size with 10% is equipped with 0.1 ton of fermention medium to three grades of seeds accesses is 0.15 ton a first class seed pot.Culture condition is: 26 ℃ of culture temperature, and 150 rev/mins of stirring velocitys, ventilation (V/V) 1: 0.4, tank pressure 0.06Mpa cultivated 86 hours;
(6) the secondary seed jar is cultivated: is in 1.5 tons of secondary seed jars with the bacterium liquid of first class seed pot with 10% the inoculum size cubic capacity that 1.0 tons of fermention mediums are housed of transferring, culture condition is: 26 ℃ of culture temperature, 130 rev/mins of stirring velocitys, ventilation (V/V) 1: 0.5, tank pressure 0.06Mpa cultivated 96 hours;
(7) fermentor cultivation: it is in 15 tons the fermentor tank that the inoculum size with 10% inserts the cubic capacity that 10 tons of fermention mediums are housed with secondary seed jar seed.Culture condition is: 26 ℃ of culture temperature, and 120 rev/mins of stirring velocitys, ventilation (V/V) 1: 0.6, tank pressure 0.08Mpa cultivated 150 hours.
Slant medium consists of the PDA slant medium in this example.
Seed culture medium and fermention medium consist of in this example: starch 3%, sucrose 3%, glucose 3%, peptone 0.5%, defatted soybean protein powder 3.0%, potassium primary phosphate 0.13%, sal epsom 0.10%, vitamins B
12PPm, PH7.0.
The dried mycelia yield of gleoesterum incarnatum is 3.0% in this example.
Claims (2)
1. gleoesterum incarnatum 10 ton liquid submerged femrentation culturing methods is characterized in that comprising the steps:
(1) slant strains activation culture: the gleoesterum incarnatum slant strains of preservation is transferred on the slant medium, cultivated 96～120 hours, and covered with the inclined-plane to mycelia and get final product for 22～28 ℃;
(2) first order seed is cultivated: access of above-mentioned gleoesterum incarnatum slant strains picking is equipped with carries out first order seed in 500 ml shake flasks of 100 milliliters of substratum and cultivate culture condition: 80～180 rev/mins of rotary shaking tables, cultivated 96～130 hours for 22～28 ℃;
(3) secondary seed is cultivated: the one-level shake-flask seed is inserted with 5～20% inoculum sizes carry out the secondary seed cultivation in 500 ml shake flasks that 100 milliliters of substratum are housed, culture condition: 80～180 rev/mins of rotary shaking tables, cultivated 96～130 hours for 22～28 ℃;
(4) three grades of seed culture: the secondary shake-flask seed inserted in 5000 ml shake flasks that 1000 milliliters of substratum are housed with 5～20% inoculum sizes carry out three grades of seed culture, culture condition: 80～180 rev/mins of rotary shaking tables, cultivated 72～96 hours for 22～28 ℃;
(5) first class seed pot is cultivated: the cubic capacity that the inoculum size with 5～20% is equipped with 0.1 ton of fermention medium to three grades of seeds accesses is 0.15 ton a first class seed pot; Culture condition is: 22～28 ℃ of culture temperature, and 80～180 rev/mins of stirring velocitys, ventilation are 1: 0.3～0.8, tank pressure 0.03～0.1Mpa cultivated 72～96 hours;
(6) the secondary seed jar is cultivated: is in 1.5 tons of secondary seed jars with the bacterium liquid of first class seed pot with 5～20% the inoculum size cubic capacity that 1.0 tons of fermention mediums are housed of transferring, culture condition is: 22～28 ℃ of culture temperature, 80～180 rev/mins of stirring velocitys, ventilation is 1: 0.3～0.8, tank pressure 0.03～0.1Mpa cultivated 72～96 hours;
(7) fermentor cultivation: it is in 15 tons the fermentor tank that the inoculum size with 5～20% inserts the cubic capacity that 10 tons of fermention mediums are housed with secondary seed jar seed; Culture condition is: 22～28 ℃ of culture temperature, and 80～180 rev/mins of stirring velocitys, ventilation are 1: 0.3～0.8, tank pressure 0.03～0.1Mpa cultivated 150～200 hours;
Described seed is identical with the fermention medium composition, and its ratio is: starch 2～3%, sucrose 3～5%, glucose 1～3%, peptone 0.5%, defatted soybean protein powder 2～3%, potassium primary phosphate 0.13～0.2%, sal epsom 0.1～0.15%, vitamins B
12PPm, PH6～7.
2. according to described a kind of gleoesterum incarnatum 10 ton liquid submerged femrentation culturing methods of claim 1, wherein culture temperature is 25 ℃.
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| CNB2004100690639A CN100340669C (en) | 2004-07-19 | 2004-07-19 | 10 ton liquid submerged femrentation culturing method of Gleoesterum incarnatum |
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| CNB2004100690639A CN100340669C (en) | 2004-07-19 | 2004-07-19 | 10 ton liquid submerged femrentation culturing method of Gleoesterum incarnatum |
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| CN100340669C true CN100340669C (en) | 2007-10-03 |
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| CN100394862C (en) * | 2006-02-22 | 2008-06-18 | 中国食品发酵工业研究院 | Elm tree mushroom feedstuff addictive, its preparation method and application |
| CN102550293B (en) * | 2012-02-03 | 2014-05-21 | 连云港市农业科学院 | Method for liquid fermentation cultivation of Agaricus bisporus strain |
| CN102550294B (en) * | 2012-02-03 | 2014-05-21 | 连云港市农业科学院 | Method for liquid fermentation cultivation of Pleurotus cornucopiae strain |
| CN103081723A (en) * | 2013-02-17 | 2013-05-08 | 哈尔滨伟平科技开发有限公司 | Preparation method for Gloeostereum incarnatum powder |
| CN109825445B (en) * | 2019-04-02 | 2020-10-30 | 鲁东大学 | Elm fungus liquid fermentation medium and fermentation culture method |
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Non-Patent Citations (1)
| Title |
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| 胶韧革菌Gloeostereumincarnatum S.ItoetⅠ mai研究Ⅱ──深层培养 太龙杰，栾恒淳，赵强，李治平，东北师大学学报自然科学版，第3期 1997 * |
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